Presenilin 1 and 2 (PS1, PS2) both undergo regulated endoproteolytic processing to yield two stable fragments. We have recently discovered that PS1 and PS2 will be alternatively cleaved at sites distal to the normal cleavage sites during apoptosis, and when over-expressed in transfected cell lines. Apoptosis-associated endoproteolysis of PS1 and PS2 can be blocked by the apoptotic cysteine protease inhibitors, zVAD, a general caspase inhibitor, and zDEVD, a selective caspase-3 (CPP32) inhibitor, indicating that the enzyme responsible is a caspase-3 family protease. We have identified the apoptotic cleavage site in PS2 as DSYDS (a.a. 326-330) by site directed mutagenesis. In support of a potential role for apoptotic cleavage of the presenilins in FAD, the ratio of apoptotic:normal cleavage fragments was elevated by over 3-fold in cells expressing the FAD mutant PS2-N141I as compared to w.t. PS2-expressing cells. Since FAD mutations in PS1/PS2 have been associated with an elevated ration of Abeta42:Abeta40, we have generated preliminary data to test whether inhibition of the caspase-3 mediated cleavage of mutant PS2 could repress the increased Abeta42:Abeta40 ration associated with FAD mutations. Treatment with zVAD repressed the increases in Abeta42:total Abeta ratio associated with the N141I and M239V FAD mutations in PS2 by 44%. Collectively, these findings not only suggest that the presenilins may be cell death substrates, but not apoptotic cleavage of the presenilins by a caspase 3 family protease may alter the Abeta42:Abeta40 ration and play a role in the pathogenesis of FAD. In view of these novel findings, we will test the following hypotheses: 1. Presenilin FAD mutations lead to increased apoptosis- associated endoproteolysis of the presenilins by a caspase-3 family protease; 2. Apoptosis-associated cleavage of the presenilins leads to an increased Abeta42:Abeta40 ratio; and 4. Apoptotic endoproteolysis of the presenilins alters the subcellular distribution and/or molecular interactions of the presenilins.